Micro- and nanoparticles can be divided into three large groups according to their composition, namely particles composed of:    I. pure drug,    II. pure matrix material (e.g. polymers, natural macromolecules, lipids),    III. matrix material loaded with active ingredient.
Particle sizes over 10 μm are easily accessible by conventional size reduction techniques, e.g. grinding with a pestle, optionally accompanied by nitrogen cooling. It is more difficult to prepare superfine particles smaller than 10-20 μm and in particular nanoparticles smaller than 1 μm, in particular in the range of a few 100 nm.
Air-jet milling gives particle distributions of up to 25 μm (Peters, K., Nanosuspensions for the i.v. administration of poorly soluble drugs—stability during sterilization and long-term storage, 22nd Int. Symp. CRS, 1995, 2212); in addition, the thermal load and exposure to oxygen can impair the chemical stability of sensitive active ingredients.
Although wet-grinding processes (List, P. H., Arzneiformenlehre, 3rd Edition, 1982, WVG, Stuttgart) in water reduce the temperature load upon suitable cooling, they are unsuitable for hydrolysis-sensitive active ingredients.
An alternative preparation process is the precipitation of the particles, e.g. for the preparation of drug nanoparticles (so-called hydrosols) (Sucker, H., Hydrosole-eine Alternative far die parenterale Anwendung von schwer wasserlöslichen Wirkstoffen, in: Müller, R. H., Hildebrand, G. E., (Eds.), Pharmazeutische Technologie: Moderne Arzneiformen, 2nd Edition, 1998, WVG, Stuttgart). A disadvantage here is that organic solvents must be used as a rule (residue in the product). A further problem is that the drug must at least be soluble in a solvent. At the same time, this solvent must also be miscible with a non-solvent in order to precipitate out the particles by the addition of the solvent to the non-solvent according to Ostwald-Mier. The resulting particles must then be prevented from growing during the precipitation process by skilful selection of the stabilizing surfactant mixture and be stabilized for long-term storage.
Other processes for preparing micro- and nanoparticles are e.g. spray-drying (Wagenaar, B. W., Müller, B. W., Piroxicam release from spray-dried biodegradable microspheres, Biomaterials 1994, 15, 49-53), solvent evaporation methods (Nihant, N., et al, Polylactide Microparticles Prepared by Double Emulsion/Evaporation Technique. I. Effect of Primary Emulsion Stability, Pharm. Res., 1994, 11, 1479-1484), solvent deposition and phase separation (Speiser, P. P., Nanopartikel, in: Müller, R. H., Hildebrand, G. E., (Eds.), Pharmazeutische Technologie: Moderne Arzneiformen, 2nd edition, 1998, WVG, Stuttgart, 339-357). However, all contain organic solvents as a rule, and in addition, contact with water is unavoidable (Fahr, A. Kissel, T., Mikropartikel and Implantate: Arzneiformen zur parenteralen Applikation, in: Müller, R. H., Hildebrand, G. E., (Eds.), Pharmazeutische Technologie Moderne Arzneiformen, 2nd Edition, 1998, WVG, Stuttgart, 243-259).
As an alternative process for preparing micro- and nanoparticles via particle reduction whilst avoiding organic, toxicologically problematical solvents, high-pressure homogenization was then used. The polymer to be reduced (Müller, B. W., Verfahren zur Herstellung von Pseudolatices und Mikro-oder Nanopartikeln und diese enthaltenden pharmazeutischen Prdparaten, EP 0 605 933 B1, 1998) or drug (Liversidge, G. G. Surface-modified drug nanoparticles, U.S. Pat. No. 5,145,684, 1991; Haynes, D. H., Phospholipid-coated microcrystals: injectable formulations of water-insoluble drugs, U.S. Pat. No. 5,091,187, 1992; Westesen, K., Solid lipid particles, particles of bioactive agents and methods for the manufacture and use thereof, International Patent Application WO 94/20072, 1994) is dispersed in water and the suspension then passed through the high-pressure homogenizer. A disadvantage here is that in the case of all processes, the particles to be reduced are exposed to water. In particular, it is stated that, in the case of polymers, the temperature load is also to be raised and possibly a toxicologically undesirable plasticizer must be added, e.g. 0.3-10% in the case of ethyl cellulose (Müller, B. W., Verfahren zur Herstellung von Pseudolatices und Mikro-oder Nanopartikeln und diese enthaltenden pharmazeutischen Präparaten, EP 0 605 933 B1, 1998). Drugs are also melted (Westesen, K., Solid lipid particles, particles of bioactive agents and methods for the manufacture and use thereof, International Patent Application WO 94/20072, 1994) which, in addition to the impairment of chemical stability, also tend not to crystallize again after homogenization (Siekmann, B., Westesen, K., Preparation and physicochemical characterization of aqueous dispersions of coenzyme Q10 nanoparticles, Pharm. Res., 1995, 12, 201-208).
Thus in general, for a gentler size reduction process, depending on the properties of the material to be homogenized, it is necessary:                to minimize or exclude contact with water        to exclude the use of toxicologically undesirable organic solvents such as dichloromethane        to minimize or avoid the temperature load        to avoid the addition of toxicologically undesirable additives such as plasticizers        to minimize or exclude exposure to oxygen        to avoid melting and to keep the substances to be processed in solid state.        
The present invention realizes a gentle reduction process by homogenization in which, depending on the properties of the substance to be processed, one or more or all of these parameters are fulfilled simultaneously. If the meeting of a parameter is not essential (e.g. exclusion of oxygen is not necessary), then it waived avoided on economic grounds in order to make the process as economical as possible.
The reduction principle of high-pressure homogenization is cavitation (Müller, R. H., Böhm, B. H. L., Grau, M. J., Nanosuspensions—Formulierungen für schwerlösliche Arzneistoffe mit Beringer Bioverfügbarkeit: I. Herstellung and Eigenschaften, Pharm. Ind., 1999, 74-78). Water boils when the static pressure acting on it (e.g. air pressure) is equal to or less than the vapour pressure. In the high-pressure homogenizer, liquid flows at a very high speed so that the static pressure sinks below the vapour pressure of water, this is transformed into the gaseous state and forms gas bubbles. When the gas bubbles collapse (e.g. on leaving the homogenization gap), this implosion leads to strong shock waves which lead to particle reduction. The reduction of substances by high-pressure homogenization was therefore previously carried out in water and not in liquids with a lower vapour pressure. Even high-pressure homogenization at increased temperature is recommended load (well above room temperature, e.g. at 60-90° C.) as the difference between static pressure (e.g. in homogenization gap) and vapour pressure of the water can then be more easily overcome. In particular, homogenization was not carried out at lower temperatures as, because the vapour pressure of the water is less at lower temperatures, the difference between static pressure and vapour pressure increases and no cavitation occurs. In particular when reducing polymers, even a temperature increase is described as insufficient for an effective reduction, and plasticizers must be added to the polymers (Müller, B. W., Verfahren zur Herstellung von Pseudolatices und Mikro-oder Nanopartikeln und diese enthaltenden pharmazeutischen Präparaten, EP 0 605 933 B1, 1998).
In the invention, it is not water but non-aqueous liquids, in particular also with a lower vapour pressure (liquid polyethylene glycols, anhydrous glycerine) that are used in the homogenization process. Surprisingly, it was shown that superfine microparticles and nanoparticles could also be prepared thereby (examples 1-6). Compared with particles which were homogenized in water, negligible differences resulted (example 3). Homogenization in anhydrous media was carried out for pure active ingredients (e.g. drugs, cosmetic active ingredients, etc.), synthetic polymers and natural macromolecules as well as for active-ingredient-charged polymers.
Depending on the degree of hydrolysis sensitivity of active ingredients, small proportions of water are tolerated in the dispersion medium. Thus proportions of water were added to the dispersion medium in order to improve the uniformity of the particle dispersion (example 7). The average diameter of the particle dispersion shows little change compared with anhydrous dispersion medium (example 6). However, the 95% diameter sinks slightly, which is an indication of the presence of a few larger particles in addition to the main population of the particles (example 13). Irrespective of this, certain proportions of water are often desired in the further processing of the particle dispersion (e.g. in PEG 400 upon packing in soft gelatine capsules, the PEG should contain a certain proportion of moisturiser so that no water is removed from the gelatine capsule wall itself and the capsule thereby becomes brittle). A condition for this is however at least a low solubility of water in the dispersion medium or miscibility. Added water proportions were e.g. 1%, 5% and 10% (e.g. example 7). Surprisingly, these water proportions—contrary to the theoretical considerations—had no reduction-promoting influence (little change in 50% diameter).
Higher proportions of water were also used (the maximum quantities of water used were 80% or 99%), the particle size decreasing insubstantially or not at all compared with the anhydrous medium (e.g. examples 7 and 8). For most products, such minimal differences are irrelevant to product quality. For suspensions for intravenous injection, it is irrelevant to the avoidance of capillary blockage whether the average diameter is 0.6 μm or 0.7 μm as long as it remains clearly below the smallest size of capillaries of 5-6 μm for the avoidance of capillary blockage (embolism). These results confirm that an external water phase is not necessary to achieve a product of sufficient fineness.
The proportion of microparticles with a size clearly above the average 50% diameter is a function of the number of homogenization cycles. It decreases (i.e. the D95% or D90% as a measure of this proportion decreases) as the number of cycles increases (example 13). To reduce the proportion of microparticles—e.g. in view of i.v. application—the number of cycles can generally be increased so that an addition of water to the dispersion medium is not necessary for this either.
An addition of water which does not impair the stability of active ingredients is also advisable if substances or polymers are dissolved in this water which are not, or not sufficiently, soluble in the anhydrous solvent, but are desirable for the final formulation. Examples are hydroxypropyl methylcellulose (HPMC) as a structuring excipient or PEG 6000 as mould release agent if the micro- or nanoparticle dispersion is to be converted into a dry formulation such as a tablet or pellet. Gelation agents, e.g. miglyol gel (solution of Aerosil with low water content to promote gelation in oil via hydroxyl groups of the water) are also advisable.
To examine the influence of a plasticizer, in a comparable manner to Müller, B. W., Verfahren zur Herstellung von Pseudolatices und Mikro-oder Nanopartikeln und diese enthaltenden pharmazeutischen Präparaten, EP 0 605 933 B1, 1998, ethyl cellulose with an added 1.74% (m/m relative to the polymer) plasticizer was homogenized at increased temperature and compared with a microparticle suspension prepared without added plasticizer (example 9). The differences in the particle sizes were small or the plasticizer-free dispersion surprisingly even showed smaller particle sizes, so that toxicologically undesired plasticizers can be dispensed with—contrary to expectations on the basis of the literature.
For polymers such as ethyl cellulose (Müller, B. W., Verfahren zur Herstellung von Pseudolatices und Mikro-oder Nanopartikeln und diese enthaltenden pharmazeutischen Präparaten, EP 0 605 933 B1, 1998), homogenization at higher temperatures should lead to smaller particles. This is based on the theoretical considerations that the difference between static pressure in the homogenizer and the vapour pressure of the dispersion medium is smaller and the softening point of the polymers is approached. Ethyl cellulose was therefore homogenized at different temperatures and the particle sizes compared (example 10). The differences were minimal and as a rule not relevant for the product quality. Thus operation is also possible for these substances at 40-60° C. or slightly above or at room temperature (20° C.) instead of 85° C. without loss of product-relevant quality or particle size.
High-pressure homogenization involves the dissipation of flow energy in heat (Jahnke, S., Theorie der Hochdruckhomogenisation, Workshop Dispergiertechnik, 4th Expert Meeting, cdc 1999), the product warms up (e.g. per cycle by approx. 10-20° C. in the case of LAB 40, APV Deutschland GmbH, Lubeck, Germany). For very temperature-sensitive substances, removal of this heat from the product should not wait until the product container stage but preferably already take place beforehand in the homogenization tower during the reduction process. In these cases, the process is carried out at lower temperature (example 14), i.e. with cooling at 4° C. or else well below 0° C., e.g. at −20° C. or −50° C., which is only possible as a purely external phase avoiding water. Contrary to theoretical considerations (even lower vapour pressure of water at these low temperatures), the high pressure homogenization was, surprisingly, sufficiently effective for preparing superfine particle dispersions. Further measures are degassing of the dispersion medium (e.g. in a vacuum or by heating) and additionally protective gassing (e.g. with nitrogen) (example 16).